Download (dendritic) cells

Dendritic Cell Therapy
The Immune System
The immune system is the body's defense system. It works on three different
levels. The first level is the anatomic response. It consists of anatomical barriers
to foreign particles and includes the skin and acid in the stomach. Anatomic
barriers prevent foreign substances from entering the body. If foreign particles
pass through the first line of defense the second line of defense called the
inflammatory response kicks in. The third line of defense is the immune
response. It is the main player in specific immune defense.
The cells of the immune system mount the immune response. These cells are
also called white blood cells.
There are several types: The neutrophils are responsible for killing bacteria and
yeast and are the first white blood cells at the site of an infection. The eosinophils
play a part in delayed reactions to foreigners. The key players that will be
discussed here are the monocytes and the lymphocytes. Monocytes are
scavangers. They scour the body for anything out of place. They can engulf
foreign particles and chew off pieces of tumor cells. Lymphocytes are not able to
engulf any foreign particles or eat tumor cells. They take the information given to
them by monocytes and monocyte-like cells and do their job. There are several
types of lymphocytes. The types essential to this topic are the B lymphocytes,
and the T lymphocytes. The B lymphocytes get their characteristics after being
nurtured in the bone marrow, hence the B. B lymphocytes are primarily
responsible for producing antibodies. Antibodies can inactive bacteria, fungi, and
viruses and make them and other foreign particles easier to see by the rest of the
immune system. T lymphocytes mature in the thymus gland, which is located
under the breast bone, hence the T. For the purposes of this topic they can be
divided into three major categories: the T helper cells, the T suppressor cells,
and the cytotoxic T cells. The T helper and suppressor cells do exactly what their
names imply. The cytotoxic T cells are primarily responsible for killing virally
infected, and tumor cells.
The Immune System and Cancer
In order for cancer to occur, the immune system must have failed. The normal
sequence of events when the immune system comes across tumor cells follows.
An immune cell called the macrophage (also called a monocyte) comes into
contact with a cancerous or precancerous cell. This cell has some strange
surface features. The strange features signal the macrophage that the cell is not
healthy and that the macrophage should take a bite out of it.
The macrophage then begins to digest the bite of the tumor cell. Several little
packets of enzymes act like a cellular stomach and break down the piece into
smaller and smaller pieces.
There are two possible scenarios that can happen next. The macrophage can
either hand off these little pieces of tumor cell to another type of immune cell, or it
can transform itself into another, specialized immune cell called a dendritic cell.
There is more and more evidence that the latter happens most often.
Dendritic cells are found in all tissues of the body, and many of them began as
macrophages. The first dendritic cell discovered is found throughout the skin and
is called a Langerhan's cell.
Now that the macrophage has digested pieces of the tumor cell, it transforms into
the dendritic cell. The dendritic cell is a much more effective messenger. When it
is fully mature, it gives the information about the tumor contained in the small
digested packets to the rest of the immune system. A key point here is that the
dendritic cell must be mature to effectively present the tumor information. The
cell needs to have additional markers on its surface that the other immune cells
can recognize. These markers are called co-stimulatory molecules and are
shown as white crosses on the picture of the mature dendritic cell below.
When the dendritic cell begins to mature, it also starts moving, or migrating
toward a lymph node. The lymph nodes contain large numbers of lymphocytes,
another type of immune cell. Everyone probably remembers having enlarged
lymph nodes in their neck when they had a sore throat. The lymph nodes are
where the action is when it comes to the immune system. There are areas in the
body that contain large numbers of lymph nodes. The neck, armpits, and groin
areas all have clusters of nodes that lie close to the skin.
So the mature dendritic cell has migrated to the lymph node. There it comes in
contact with different kinds of lymphocytes. If it has matured properly, the costimulatory molecules on its surface will help pass the tumor information along to
the cytotoxic T lymphocytes, or CTLs. The CTLs are the body's main defense
against tumor cells. When the right CTL comes in contact with the dendritic cell, it
will become activated and begin to divide, effectively making an army of cloned
soldiers ready to kill any cancerous or pre-cancerous cell having the same
altered membrane discovered by the macrophage.
When the CTL soldiers come in contact with cells that have the same surface as
the original cancerous cell, they bind to it. They then release a chemical that
pokes tiny holes in the membrane of the tumor cell, and the tumor cell spills its
guts and dies.
Let's summarize what happens normally in the body after a normal cell turns
cancerous. First, a macrophage comes in contact with the tumor cell, which has
a different type of membrane that signals the macrophage to eat part of it. The
macrophage then digests the eaten tumor cell fragment and starts to turn into a
dendritic cell. It then begins to mature, and travels to a nearby lymph node and
hands off the tumor cell information to CTLs. The CTLs then divide, circulate
throughout the body, and kill any tumor cells they come in contact with.
Above we covered what happens normally in the body when a cell becomes
cancerous. This process occurs countless times as cells get genetic mutations
and become cancerous. But, if you have cancer, then something must have gone
wrong. Did the macrophage fail to recognize the funny cell surface? Did
macrophages not become dendritic cells? Or did the T cells not do their job? It is
impossible to tell for sure but there are some clues that the problem is with the
dendritic cells.
Lately several research groups have been looking at the dendritic cells in and
around tumors. What they're finding is that there are dendritic cells there, but
they are immature. They don't have the co-stimulatory molecules necessary for
the successful hand off of the tumor cell membrane information to the T cells.
Moreover, because they are immature, they are much less likely to migrate to the
lymph nodes to make the hand off.
To make a football analogy, the dendritic cell is the quarterback and needs to
hand off the football to the running back (the T cell). In order to do that, he needs
to move toward the running back and hand him the ball without fumbling. When
the dendritic cell is immature, it just stands in one place and drops the ball. If that
continues to happen, your team never scores and ultimately loses the game.
If you cut up a piece of tumor from kidney cancer or renal cell carcinoma and
look at it under the microscope, you'll find millions of dendritic cells many more
than in any other type of tumor. Expectedly, the majority of these dendritic cells
are immature they don't have co-stimulatory molecules on them. What makes
this more interesting is the fact that kidney cancer is the most likely type of
cancer to disappear without a trace without any treatment, or spontaneously
remiss. What I believe happens when someone has a spontaneous remission is
the conditions in and around the tumor change enough to allow at least some of
the dendritic cells to mature. This is more likely to induce a remission in renal cell
carcinoma simply because of the larger numbers of dendritic cells.
So, what can you do to get dendritic cells to hand off information about your
tumor cells to your CTLs? Both animal and human trials of using dendritic cells in
the treatment of cancer have shown promising results and give us a direction in
which to go.
Mayordomo et al.1 inoculated mice with different types of cancer and allowed the
tumors to develop for one to two weeks. Dendritic cells were isolated from the
bone marrow of these mice, cultured with some growth factors, and exposed to
tumor peptides (information about the tumor cell membranes). These primed'
dendritic cells were then injected back into the tumor-bearing mice every four to
seven days. Recovery, measured as halting of tumor growth and subsequent
regression, was seen 7-10 days after the first injection of dendritic cells. Using
this treatment, cure rates of 80% for mice with Lewis lung carcinoma and 90% for
mice with sarcoma were achieved.
In a similar study, Nair, et al.2 induced malignant melanoma lung metastases
(new tumors that spread from the first tumor) in mice, and then surgically
removed the primary tumor. The mice were then treated with dendritic cells which
had been primed' in a manner similar to that described above. Of the seven
treated animals, four had no visible lung tumors, two had fewer than five
remaining tumor nodules, and one mouse had 15 nodules. The number of
nodules in control mice, those that did not receive dendritic cell therapy, were too
many to count, but comprised approximately three-quarters of the lung by weight.
Hsu et al.3 at Stanford University pioneered the use of dendritic cell therapy of
cancer in humans. They purified dendritic cells from the circulating blood of four
patients with B cell lymphoma previously treated with chemotherapy. The
dendritic cells were cultured and treated with antigen (tumor cell membrane
information) derived from the patients' tumors. The dendritic cells were given
using vein injections on 4 occasions; subcutaneous (under the skin) injections of
the tumor antigen and a protein that helps stimulate an immune response were
injected two weeks after each dendritic cell injection. All of the patients
developed measurable T cell immune responses after one or two vaccinations.
Meaningful clinical responses were seen. There was one partial response, one
minor response, and disease stabilization in three patients with progressive
measurable disease, and a complete response in a patient with minimal
detectable disease. All of the patients have remained progression-free for two
years.
Gerald Murphy, M.D.4, and his team at Northwest Hospital's Pacific Northwest
Cancer Foundation have been testing the use of dendritic cells in patients with
advanced prostate cancer. They cultured monocytes (macrophages) from
circulating blood with growth factors and small pieces of protein found on the
surface of prostate tumor cells. These dendritic cells were then reinfused into the
patients through an intravenous drip. They performed two studies. More than
27% of study patients who participated in both clinical trials showed some
improvement and the disease was stable in another 33%. All of the patients in
the study had advanced prostate cancer and were unresponsive to conventional
therapies, including hormone treatment.
In addition to lymphoma and prostate cancer, the deadly skin cancer malignant
melanoma has been treated successfully using dendritic cell therapy. In a recent
human study by Nestle et al5, dendritic cells were used to treat sixteen patients
with advanced metastatic (the cancer has spread) melanoma. Objective
responses were seen in 5 of the 16 patients. There were two complete
responses and three partial responses with regression of metastases in several
organs, including skin, lung, and pancreas. The participants were followed for 15
months and no cases of autoimmunity a potential side effect of the therapy were
found in any of the patients. The authors concluded, vaccination with autologous
[derived from the person's own body] dendritic cells generated from peripheral
blood is a safe and promising approach in the treatment of metastatic melanoma.
In the studies quoted above, there were little to no side effects. Murphy reports
transient hypotension (temporary low blood pressure) as the only side effect
seen in his study.
Given all of this compelling evidence that dendritic cells may hold a key position
in effective, non-toxic treatments for cancer, we began studying them.
Our research to date has focused mainly on methods of:
1. Producing large numbers of dendritic cells from the circulating blood of
cancer patients;
2. Finding the source of tumor material (antigen) for each type of tumor that
will best stimulate the T cells to proliferate and kill tumor cells; and,
3. Stimulating the dendritic cells already in and around the tumor to mature,
and become better T cell stimulators.
What we have found so far is that we can produce large numbers of dendritic
cells from the circulating blood, give them tumor antigen, and mature them.
These dendritic cells in culture are able to stimulate large numbers of T cells to
become active against tumors. We are now setting out to determine if this is
possible in humans.
In order to study the value of dendritic cells and activation of dendritic cells as
anti-tumor therapies for patients with metastatic prostate cancer we are currently
performing four clinical studies described below.
1. DENDRITIC CELLS TREATED WITH PATIENT'S OWN TUMOR MATERIAL
Before beginning on this protocol, patients must first provide tumor tissue to the
laboratory for use with the dendritic cells. Arrangements are made with your
surgeon, or urologist prior to surgery for proper collection and transport of the
tumor material to the laboratory. At the laboratory, an extract of the tumor tissue
will be made, and then filtered to make it sterile.
This study uses the patient's own tumor material and own dendritic cells.
Monocytes are first harvested from the circulating blood using a specialized
machine. The machine is called an apheresis (Greek for to take out) unit. This
type of machine is used at many American Red Cross offices to harvest blood
products such as platelets (cell fragments that help blood to clot). The procedure
is relatively simple. A needle connected to sterile, one-use tubing is placed into
each arm. The tubing is connected to the machine, and approximately one cup of
blood is circulated out of one arm, through the machine, and back into the other
arm. The machine spins the blood, removes two types of white blood cells, and
returns the fluid part of the blood and all of the other blood cells to the patient.
Using this machine allows for many more white blood cells to be collected than
by simply drawing blood because the other cells are returned to the patient. The
procedure takes about two hours.
The white blood cells are then taken to the laboratory where they are grown in a
special growth broth that begins to convert them to dendritic cells. After a few
days the filtered tumor extract is added to the growth broth. The cells are then
allowed to mature for another few days.
The cells are then tested for their maturity and purity. They are then frozen in
liquid nitrogen and are ready to be reinfused.
A portion of the cells will then be suspended in sterile saline reinfused
intraveneously (by a vein in the arm) into the patient. The patient will receive an
infusion once or twice per month.
After the cells are infused, some will migrate to the lymph nodes and some will
stay in the circulation. The likelihood is high that many of the dendritic cells will
come in contact with T-lymphoctyes (CTL's) and stimulate them to divide and
recognize and kill tumor cells. Measuring the serum levels of prostate specific
antigen, or PSA can allow your doctor to follow the course of prostate cancer.
This test is used to determine if the treatment has been effective.
2. DENDRITIC CELLS TREATED WITH PURIFIED TUMOR ANTIGEN
This procedure is the same as number one with one exception. The agent used
to prime or activate the dendritic cells is not an extract of the patient's own tumor.
Instead, a known tumor antigen (molecule that has information about the outside
of tumor cells) is placed with the dendritic cells while they grow and mature in
culture. PSMA, or prostate specific membrane antigen, is used.
PSMA is found primarily on the outside prostate cells. The priming agents used
by Murphy, described above, were fragments of the PSMA molecule.
Measurement of the serum levels of PSA will also be used to determine if the
treatment has been effective.
3. STIMULATION OF IMMATURE DENDRITIC CELLS TO BECOME MATURE
DENDRITIC CELLS.
In many types of tumors the dendritic cells in and around the tumors are
immature meaning they are there, they just can not effectively pass along
information about the tumor cells to the rest of the immune system, in particular
to the CTL's, or T-lymphocytes.
One way to convert immature dendritic cells into mature ones is by using a
mixture of growth factors and stimulating factors called cytokines. Many people
have heard of some of these cytokines. Interleukin 2, interferon, and tumor
necrosis factor are a few. These growth factors are found naturally in the body
and can be manufactured using a patient's own white blood cells. The
monocytes, when mixed with a medication made from bacterial cell walls, can
produce an effective mixture of cytokines. The monocytes are mixed with the
medicine and grown in an incubator for 2 days. Then the culture medium, or
broth, is collected. It contains a lot of cytokines. We call this mixture MCM, which
stands for monocyte-conditioned-medium. When MCM is placed with immature
dendritic cells, a majority of them become mature. They can then migrate and
effectively convey information about the tumor to lymphocytes, stimulating them
to divide at the same time.
Because the MCM is prepared from the patient's own cells, the dosage varies
from individual to individual. Because these cytokines are the same molecules
that can make you feel poorly and give you a fever when you have the flu, the
dosage is increased gradually to make sure the patient doesn't experience those
side effects. The MCM is sterile filtered, and then mixed with sterile saline. It is
given in the vein the same as the dendritic cells. An increased dose will be given
daily until a rise in the patient's body temperature is seen. That dose will then be
given daily for two weeks.
The serum PSA test is also used to follow the effects of the treatment on the
course of prostate cancer.
4. CELL FRAGMENTS OF DENDRITIC CELLS PRIMED WITH TUMOR CELL
ANTIGENS
Researchers in France recently discovered that when dendritic cells are grown in
culture they form and release small cell fragments named exosomes. These
exosomes contain all of the information necessary to activate cytotoxic T
lymphocytes against tumor cells. In one study, a significant number of tumorbearing animals had a complete remission after receiving a single injection of
exosomes into the skin.
This study begins similarly to studies one and two. Monocytes are harvested
from the peripheral blood. They are cultured with cytokines and tumor cell
antigens are added. The cells are then allowed to mature. Instead of using the
dendritic cells the liquid in which the cells are growing is collected. The
exosomes are then removed from the liquid. They are sterile filtered and injected
into the skin just above the lymph glands in the groin. The theory is that there
they interact with the dendritic cells in the skin, which move into the lymph glands
and present tumor antigen to the lymphocytes. Which then divide, circulate in the
body, and kill the tumor cells they come in contact with.
Source: www.immunemedicine.com